Waterborne pathogens like Vibrio cholerae pose significant threats to global health. V. cholerae can persist in the aquatic environment, and it can emerge to cause devastating cholera outbreaks in endemic regions and in vulnerable areas where infrastructure has been compromised and populations have been displaced. The host- pathogen interactions that dictate disease outcome and cholera transmission dynamics occur in the context of a complex microbial ecosystem that includes predatory bacterial viruses (phages). Knowledge of the molecular consequences of phage predation on the selection of epidemic strains is needed to enhance our understanding of the forces impacting the evolution and epidemiology of V. cholerae. We found that a novel group of mobile genetic elements called PLEs (phage-inducible chromosomal island-like elements) are a key weapon that epidemic V. cholerae uses to protect against phage attack. Our hypothesis is that PLEs are potent, highly specific, antiviral barriers that act through two distinct mechanisms to ensure that phage do not propagate and spread to neighboring V. cholerae cells. Our data indicate that PLEs act by inhibiting the production of essential phage proteins and by rupturing the infected bacterial cell before all progeny phage complete their replication cycle. To understand PLE activity in mechanistic detail we will pursue the following specific aims: 1) We will define the regulatory network that permits the rapid stimulation of PLE activity when V. cholerae is under attack by phage and determine how specificity is imparted onto PLE-phage interactions. 2) We will determine how PLE interferes with the ability of phage to produce essential proteins. Under this aim we will use a combination of functional genomic approaches and mutational analyses to separate this aspect of PLE activity from PLE-mediated cell lysis. 3) We will determine the molecular basis for PLE-mediated bacterial cell lysis. We will characterize suppressors that are insensitive to cytotoxic PLE-encoded products to understand how PLE efficiently kills phage-infected V. cholerae. The proposed studies will establish how phage predation shapes the genome of epidemic V. cholerae. This knowledge will enhance our understanding of phage-mediated perturbations to microbial populations in healthy and diseased states, and further our ability to manipulate these communities for therapeutic or prophylactic benefit.